Risk factors associated with dengue complications and death: A cohort study in Peru

Background Dengue has emerged as an unprecedented epidemic in Peru, and it is anticipated that this issue will escalate further owing to climate change. This study aimed to determine the risk factors associated with death from dengue in patients treated at Hospital II in Pucallpa, Peru. Methodology This retrospective cohort study collected information from the medical records of patients with a diagnosis of dengue treated at Hospital II Pucallpa-Peru between January 2019 and March 2023. The primary outcome was death, and the secondary outcome was death, development of severe dengue, or Intensive Care Unit (ICU) admission. Cox regression models were used to determine risk factors. Findings The clinical records of 152 patients were evaluated, with a median age of 27.5 years (interquartile range, 11–45). Among all patients, 29 (19.1%) developed severe dengue, 31 (20.4%) were admitted to the ICU, and 13 (8.6%) died during follow-up. In the survival analysis, bilirubin >1.2 mg/dL was associated with a higher risk of death aHR: 11.38 (95% CI: 1.2 106.8). Additionally, factors associated with poor prognosis included having 1 to 3 comorbidities aRR: 1.92 (1.2 to 3.2), AST ≥251 U/L aRR: 6.79 (2.2 to 21.4), history of previous dengue aRR: 1.84 (1.0 to 3.3), and fibrinogen ≥400 mg/dL aRR: 2.23 (1.2 to 4.1). Significance Elevated bilirubin was associated with death from dengue, whereas an increase in comorbidities and a history of previous dengue were related to a poor prognosis of the disease. Early identification of severe dengue would be more feasible with improved access to laboratory testing, particularly in tropical areas with a high dengue incidence.


Introduction
Dengue has left a wave of deaths never seen before in Peru [1].Dengue is an acute disease transmitted mainly by Aedes Aegypti mosquitoes [2] in tropical and subtropical regions worldwide [3].The incidence of dengue has dramatically increased worldwide.According to the World Health Organization, an estimated 100-400 million infections occur annually.The disease is endemic in more than 100 countries, including the Americas, which reported 3.1 million cases in 2019, with more than 25 000 classifieds as severe and the largest number of dengue cases ever reported [4].
Peru is a dengue endemic country [5], where more than half of the population is at risk of dengue infection due to 17 of the 25 departments believed to be inhabited by mosquito vectors [6].In recent years, Peru has faced record numbers of dengue cases, particularly in the Amazon region, which are likely associated with climate change [5].This is particularly important as Peru experiences an unprecedented heatwave [7], surpassing the historical maximum of 41.1˚C in 1963 [8].This increase in temperature owing to climate change could negatively influence the risk of dengue transmission [9].This highlights the need for current studies on dengue and other diseases associated with climate change [10].
Although most dengue virus infections are asymptomatic, a wide spectrum of manifestations can occur, ranging from unnoticed infections to severe cases and fatal outcomes [11].The severe form of the disease is closely associated with significant morbidity and mortality, as reflected in the alarming figure of over 10,000 estimated dengue deaths in 2013 [2,12].Risk factors such as extreme age, previous dengue infections, and the specific serotype of the virus have been linked to the development of severe cases [13,14].
Two recent systematic reviews have highlighted potential biomarkers associated with an unfavorable prognosis in dengue cases, including elevated levels of C-reactive protein, aspartate aminotransferase, and interleukin 8 and reduced levels of albumin and platelets.However, despite the substantial number of studies included in both reviews, it is important to note that they lacked representation of the Peruvian population, with less than 30% of the included studies conducted in Latin American countries [15,16], demonstrating limited evidence of dengue in Latin America, especially in Peru.Due to the alarming increase in dengue cases in Peru, it is essential to identify markers of poor prognosis for the early detection of patients with poor outcomes.This study aimed to determine the risk factors associated with death and complications of dengue in patients treated at a hospital in Peru.

Study design
We conducted a retrospective cohort study by reviewing the medical records of patients from Hospital II Pucallpa EsSalud from 2019 to 2023.This hospital belongs to the healthcare network of Ucayali and was specifically chosen because of its substantial patient volume, covering 164,318 insured individuals, and its prominence in the Peruvian Amazon as the healthcare facility in Pucallpa, the region's second most populated city [17].Our research methodology and manuscript preparation strictly adhered to the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) guidelines [18].

Population
For convenience, we performed nonprobabilistic sampling.This method was chosen because we did not have information on all patients who developed severe dengue; therefore, we decided to include all those we identified.Patients with a confirmed diagnosis of dengue were included and identified either by ns1 antigen tests or serological tests (ELISA IgM), depending on the time of illness in which the patient was.

Data collection and variable definition
We included patients hospitalized between January 1, 2019, and March 1, 2023.We selected this period because, starting in 2019, the medical records at Hospital II Pucallpa EsSalud became digital, allowing for improved patient information retrieval and collection.Patient data were accessed between July 1 and October 30, 2023.
Electronic medical records were retrieved using the Health Intelligence Service, a digital system that archives patient histories, including background information, auxiliary examinations, diagnoses, disease progression, and treatment details.This systematic approach ensured a thorough and consistent data-collection process in our study [20].
Data collection was conducted by two researchers, J.V.C. and G.T.S., who implemented an independent double-data recording system.A third researcher (C.C.C.) reviewed and compared the databases generated by J.V.C. and G.T.S. to ensure equality.When discrepancies were identified, the electronic medical records were reviewed to rectify any errors.
In addition, follow-up of deaths due to dengue was performed during the hospital stay; we considered time zero as admission to the hospital and the final time as the occurrence of death or hospital discharge of the participants.

Outcome variables
Two outcomes were considered: in-hospital mortality and dengue complications.Hospital mortality was the primary outcome variable.These data were compiled based on the outcomes documented in patients' medical records throughout the follow-up period.
As a secondary outcome, we assessed dengue complications, which, for the purposes of this study, were defined by the research team as any of the following events: admission to the Intensive Care Unit (ICU), severe dengue or death.These criteria were chosen to comprehensively capture the poor outcomes associated with dengue infections.

Statistical analysis
We described the variables using frequencies, percentages, and measures of central tendency and dispersion, as appropriate.To identify the factors linked to mortality, we initially applied Cox proportional regression models to each variable.Statistically significant variables (P <0.05) were included in the multivariate model.Similarly, to identify the characteristics associated with disease complications, we utilized Poisson regression with robust variance, incorporating only the multivariate model those variables that were statistically significant (p<0.05) in the raw regression.In both regression models, we decided not to include the urea variable in the multivariate analysis because it had a direct relationship with creatinine (both associated with kidney injury).Therefore, we prioritized the inclusion of the latter.Furthermore, we conducted survival analysis using the Kaplan-Meier method, with the log-rank test deployed to assess differences in survival functions.All analyses were performed using the statistical software Stata v17.

Ethics
This research project was approved by the Research Ethics Committee of Hospital II Pucallpa EsSalud, as evidenced by the verification code CA-04-CEI-DRAUC-ESSALUD-2023.In adherence to strict confidentiality protocols, all patient data extracted from medical records were anonymized, with random codes assigned to each record for analysis.Given the observational and retrospective nature of our study, informed consent was not sought in line with the ethical guidelines for such research methodologies.

Results
During the study period, information was collected from 163 medical records.Of these, 12 were excluded because they did not have a confirmed diagnosis of dengue.Finally, 152 medical records of patients with dengue were analyzed (Fig 1).

Factors associated with death from dengue
In multivariate analysis, bilirubin �1.2 mg/dL was associated with a higher risk of mortality (aHR:11.38;IC95%: 1.2 to 106.8) (Table 2).Fig 2 shows the survival function using Kaplan-Meier curves for the variable that was associated with the adjusted regression analysis.

Discussion
To the best of our knowledge, this is the first study in Peru to identify risk factors associated with dengue mortality and complications.A higher number of comorbidities, a history of previous dengue, and liver injury, as indicated by elevated bilirubin and transaminase levels, were the main markers associated with poor disease prognosis.
In fatal cases of dengue, the liver is the most frequently affected organ [25].Its clinical presentation can be asymptomatic and progress to acute liver failure (ALF) [26].Although its initial presentation is more commonly associated with elevated transaminase levels, elevated  bilirubin is a better marker of ALF [27,28], In our study, it was directly associated with patient mortality, like the findings reported in studies conducted in India [29] and Thailand [30].In patients with dengue, ALF is a serious complication as it can exacerbate coagulation disorders, including disseminated intravascular coagulation, infection (septicemia), renal failure, increased intracranial pressure leading to cerebral edema, and ultimately cardiopulmonary collapse, resulting in multiple organ failure and patient death [31,32].Although the pathophysiology of hepatic involvement is not fully understood, it is postulated that hepatic manifestations result from direct viral toxicity or a dysregulated immune response to the virus [25].This is supported by studies that, when evaluating patients who died from dengue, detected dengue virus RNA and the presence of viral antigens in sinusoidal endothelial cells and Kupffer cells of the liver [25,33].These findings may explain the hepatic damage observed in patients with dengue.
We found that a higher number of comorbidities was associated with an unfavorable prognosis for dengue.This could be attributed to the fact that the presence of comorbidities complicates the clinical management of patients with dengue, especially in the case of cardiovascular, chronic renal, and pulmonary diseases [34].Unlike other studies, we did not find that advanced age was associated with a higher death rate from dengue [34][35][36].This is probably due to the small number of patients included in our study, which could have reduced the power to detect differences between populations with and without dengue complications.
We observed that elevation of fibrinogen, a key player in the coagulation cascade that contributes to blood clot formation, is associated with a poor prognosis for dengue.The decrease in fibrinogen levels in severe cases of dengue is probably due to dengue infection, which mainly activates fibrinolysis in the absence of a thrombotic stimulus, leading to the direct degradation of fibrinogen and secondary activation of several procoagulant homeostatic mechanisms [37].Our results contrast with those of other studies that reported that severe forms of dengue with hemorrhagic manifestations present with prolonged clotting times and low fibrinogen levels [37][38][39][40][41][42].This discrepancy with our findings could be attributed to the fact that the increase in fibrinogen is secondary to a compensatory response to improve platelet function and reduce the risk of bleeding in patients with severe dengue [43,44].It is postulated that the development of hemorrhage in patients with dengue may be due to a combination of thrombocytopenia, coagulation imbalances, dysfunctional surviving platelets, and increased fibrinolysis [35,37].This is probably why we had very few patients with hemorrhagic manifestations of dengue in our cohort.Climate change will have multiple consequences for health worldwide [45], altering the global distribution of both infectious and zoonotic disease incidences and the risk of these diseases emerging in new regions [46]."El Niño-Southern Oscillation," the Cyclone "Yacu," and climate change likely contributed to increased vector density [47], due to the accumulation of stagnant water from heavy rains and longer warm seasons [5]; as both conditions favor mosquito proliferation [48], increasing their survival and viral replication [6].Additionally, it has been associated with bite rates [49], which in turn increases the risk of dengue transmission and other vector-borne diseases [45].To this end, serious problems of access to clean water and sanitation for rural populations and the poor management of solid waste by regional governments in Peru have been added [50].It is a combination of these circumstances that has unleashed the largest dengue epidemic in Peruvian history.
Faced with this situation, national governments of Latin America must promote intervention policies supported by scientific evidence to mitigate the probable increase in diseases related to climate change.Our study identified early laboratory markers associated with a poor disease prognosis.Providing access to laboratory tests in areas with a high prevalence of dengue could facilitate the diagnosis and identification of patients with poor prognosis, with the aim of providing them with greater surveillance and timely treatment [51].This study has certain limitations that must be considered when analyzing the results.The main limitation was its retrospective nature, which prevented the evaluation of certain variables that could be confounding factors or contribute to explaining the study phenomenon (such as self-medication and other concomitant infections).However, owing to the limited resources available, the NS1 and IgM tests for dengue are qualitative in our hospital.This limits the possibility of evaluating whether the quantitative levels of NS1 and IgM are associated with severity.Additionally, owing to the observational nature of our study, we were unable to establish causality between variables.Furthermore, the study was small and was only carried out in one institution; therefore, the results are not generalizable to the entire Peruvian population.
However, this study has important strengths.To the best of our knowledge, this is the first study to identify poor prognostic factors in patients with dengue in Peru.Furthermore, given the evident climate change and possible expansion of dengue in Latin America, especially in Peru, our data are of great relevance.As health services are often limited in resource-poor settings, any information that can help distinguish patients with severe dengue at a higher risk of progressing to death may be crucial.
In conclusion, our study revealed that liver injury, as indicated by an increase in bilirubin level, was the main predictor of mortality in patients with dengue.Additionally, a higher number of comorbidities and a history of dengue were associated with a poor prognosis.Therefore, early identification of dengue patients with poor prognosis could be improved by providing greater access to laboratory testing, particularly in tropical areas with a high prevalence of dengue.This is crucial, considering that dengue has become a significant public health problem worldwide.